Apoptosis is referred to programmed cell death in a multi-cellular organism and plays a physiological role to grow a normal cell, to maintain cell homeostasis, to prevent cancer and other diseases and to protect a living body from viral or bacterial infection.
Some Bcl-2 family proteins including Bax, Bad, Bak, Bid and Noxa are reported to stimulate apoptosis and have a Bcl-2 homology domain such as BH1, BH2, BH3 and BH4. Especially, Noxa protein is a “BH3-only” member of Bcl-2 family and first discovered in a mouse cell after exposed by X-ray. Also, it is already reported to mediate p53-activated apoptosis (Oda E. et al., Science 288 (5468): 1053˜8, 2000). BH3 is a well-known domain in Bcl-2 family and plays a critical role to induce apoptosis. It is also reported that BH3 domain derived from Bid protein may cause apoptosis by peptide itself (Letai et al., Cancer cell 2: 183˜192, 2002).
In human, it is elucidated that Noxa regulates the function of mitochondria to stimulate apoptosis. The present inventor has found a mitochondria targeting domain (hereinafter, referred to as “MTD”) at the C-terminus of Noxa protein, separate from BH3 domain acting on apoptosis (Seo et al., JBC278: 48292˜48299, 2003). In detail, Noxa protein triggers cytochrome C discharge from mitochondria, activates caspase and kills cells. MTD domain is essential to transport Noxa toward mitochondria and may lose the cell killing activity if deleted, although BH3 domain exists. Therefore, MTD domain is considered to play an important role to kill cells. Nevertheless, it is also reported that only MTD domain without BH3 domain did not bring about apoptosis.
The present inventor has identified that MTD domain, a C-terminal region of Noxa protein that is known to assist BH3 domain for targeting mitochondria, strongly killed cancer cell lines (including HeLa, PC3, A549, HEK 293, and HCT 116) when combined with R8, a protein transduction domain (hereinafter, referred to as “PTD”), tried to develop a novel cell-killing peptide (CKP) and completed the invention successfully.